Dan C. Freed

Bio: Dan C. Freed is an academic researcher from United States Military Academy. The author has contributed to research in topics: T cell & DNA vaccination. The author has an hindex of 4, co-authored 5 publications receiving 1184 citations. Previous affiliations of Dan C. Freed include Merck & Co..

Control of Viremia and Prevention of Clinical AIDS in Rhesus Monkeys by Cytokine-Augmented DNA Vaccination

Abstract: With accumulating evidence indicating the importance of cytotoxic T lymphocytes (CTLs) in containing human immunodeficiency virus-1 (HIV-1) replication in infected individuals, strategies are being pursued to elicit virus-specific CTLs with prototype HIV-1 vaccines. Here, we report the protective efficacy of vaccine-elicited immune responses against a pathogenic SHIV-89.6P challenge in rhesus monkeys. Immune responses were elicited by DNA vaccines expressing SIVmac239 Gag and HIV-1 89.6P Env, augmented by the administration of the purified fusion protein IL-2/Ig, consisting of interleukin-2 (IL-2) and the Fc portion of immunoglobulin G (IgG), or a plasmid encoding IL-2/Ig. After SHIV-89.6P infection, sham-vaccinated monkeys developed weak CTL responses, rapid loss of CD4+ T cells, no virus-specific CD4+ T cell responses, high setpoint viral loads, significant clinical disease progression, and death in half of the animals by day 140 after challenge. In contrast, all monkeys that received the DNA vaccines augmented with IL-2/Ig were infected, but demonstrated potent secondary CTL responses, stable CD4+ T cell counts, preserved virus-specific CD4+ T cell responses, low to undetectable setpoint viral loads, and no evidence of clinical disease or mortality by day 140 after challenge.

Potent CD4+ T cell responses elicited by a bicistronic HIV-1 DNA vaccine expressing gp120 and GM-CSF.

Abstract: Virus-specific CD4 + T cell responses have been shown to play a critical role in controlling HIV-1 replication. Candidate HIV-1 vaccines should therefore elicit potent CD4 + as well as CD8 + T cell responses. In this report we investigate the ability of plasmid GM-CSF to augment CD4 + T cell responses elicited by an HIV-1 gp120 DNA vaccine in mice. Coadministration of a plasmid expressing GM-CSF with the gp120 DNA vaccine led to only a marginal increase in gp120-specific splenocyte CD4 + T cell responses. However, immunization with a bicistronic plasmid that coexpressed gp120 and GM-CSF under control of a single promoter led to a dramatic augmentation of vaccine-elicited CD4 + T cell responses, as measured by both cellular proliferation and ELISPOT assays. This augmentation of CD4 + T cell responses was selective, since vaccine-elicited Ab and CD8 + T cell responses were not significantly changed by the addition of GM-CSF. A 100-fold lower dose of the gp120/GM-CSF bicistronic DNA vaccine was required to elicit detectable gp120-specific splenocyte proliferative responses compared with the monocistronic gp120 DNA vaccine. Consistent with these findings, i.m. injection of the gp120/GM-CSF bicistronic DNA vaccine evoked a more extensive cellular infiltrate at the site of inoculation than the monocistronic gp120 DNA vaccine. These results demonstrate that bicistronic DNA vaccines containing GM-CSF elicit remarkably potent CD4 + T cell responses and suggest that optimal Th cell priming requires the precise temporal and spatial codelivery of Ag and GM-CSF.

Cross-Reactivity of Anti–HIV-1 T Cell Immune Responses among the Major HIV-1 Clades in HIV-1–Positive Individuals from 4 Continents

Abstract: BACKGROUND The genetic diversity of human immunodeficiency virus type 1 (HIV-1) raises the question of whether vaccines that include a component to elicit antiviral T cell immunity based on a single viral genetic clade could provide cellular immune protection against divergent HIV-1 clades. Therefore, we quantified the cross-clade reactivity, among unvaccinated individuals, of anti-HIV-1 T cell responses to the infecting HIV-1 clade relative to other major circulating clades. METHODS Cellular immune responses to HIV-1 clades A, B, and C were compared by standardized interferon- gamma enzyme-linked immunospot assays among 250 unvaccinated individuals, infected with diverse HIV-1 clades, from Brazil, Malawi, South Africa, Thailand, and the United States. Cross-clade reactivity was evaluated by use of the ratio of responses to heterologous versus homologous (infecting) clades of HIV-1. RESULTS Cellular immune responses were predominantly focused on viral Gag and Nef proteins. Cross-clade reactivity of cellular immune responses to HIV-1 clade A, B, and C proteins was substantial for Nef proteins (ratio, 0.97 [95% confidence interval, 0.89-1.05]) and lower for Gag proteins (ratio, 0.67 [95% confidence interval, 0.62-0.73]). The difference in cross-clade reactivity to Nef and Gag proteins was significant (P<.0001). CONCLUSIONS Cross-clade reactivity of cellular immune responses can be substantial but varies by viral protein.

Cross-clade reactivity of HIV-1-specific T-cell responses in HIV-1-infected individuals from Botswana and Cameroon

Abstract: An effective HIV type 1 (HIV-1) vaccine will likely require elicitation of broadly reactive cell-mediated immune (CMI) responses against divergent HIV-1 clades. We compared anti-HIV-1 T-cell immune responses among 363 unvaccinated adults infected with diverse HIV-1 clades. Response rates to clade B Gag and/or clade B Nef in Botswana (95%) and Cameroon (98%) were similar when compared with those in countries previously studied, including Brazil (92%), Thailand (96%), South Africa (96%), Malawi (100%), and the United States (100%). Substantial cross-clade cell-mediated immune responses in Botswana and Cameroon confirm previous findings in a larger, more genetically diverse collection of HIV-1 samples.

Replication-incompetent adenoviral vaccine vector elicits effective anti-immunodeficiency-virus immunity.

Abstract: Recent studies of human immunodeficiency virus type 1 (HIV-1) infection in humans and of simian immunodeficiency virus (SIV) in rhesus monkeys have shown that resolution of the acute viral infection and control of the subsequent persistent infection are mediated by the antiviral cellular immune response. We comparatively assessed several vaccine vector delivery systems-three formulations of a plasmid DNA vector, the modified vaccinia Ankara (MVA) virus, and a replication incompetent adenovirus type 5 (Ad5) vector-expressing the SIV gag protein for their ability to elicit such immune responses in monkeys. The vaccines were tested either as a single modality or in combined modality regimens. Here we show that the most effective responses were elicited by a replication-incompetent Ad5 vector, used either alone or as a booster inoculation after priming with a DNA vector. After challenge with a pathogenic HIV-SIV hybrid virus (SHIV), the animals immunized with Ad5 vector exhibited the most pronounced attenuation of the virus infection. The replication-defective adenovirus is a promising vaccine vector for development of an HIV-1 vaccine.

Control of a mucosal challenge and prevention of AIDS by a multiprotein DNA/MVA vaccine.

Abstract: Heterologous prime/boost regimens have the potential for raising high levels of immune responses. Here we report that DNA priming followed by a recombinant modified vaccinia Ankara (rMVA) booster controlled a highly pathogenic immunodeficiency virus challenge in a rhesus macaque model. Both the DNA and rMVA components of the vaccine expressed multiple immunodeficiency virus proteins. Two DNA inoculations at 0 and 8 weeks and a single rMVA booster at 24 weeks effectively controlled an intrarectal challenge administered 7 months after the booster. These findings provide hope that a relatively simple multiprotein DNA/MVA vaccine can help to control the acquired immune deficiency syndrome epidemic.

DNA vaccines: ready for prime time?

Abstract: Since the discovery, over a decade and a half ago, that genetically engineered DNA can be delivered in vaccine form and elicit an immune response, there has been much progress in understanding the basic biology of this platform. A large amount of data has been generated in preclinical model systems, and more sustained cellular responses and more consistent antibody responses are being observed in the clinic. Four DNA vaccine products have recently been approved, all in the area of veterinary medicine. These results suggest a productive future for this technology as more optimized constructs, better trial designs and improved platforms are being brought into the clinic.

Evidence of HIV-1 Adaptation to HLA-Restricted Immune Responses at a Population Level

Abstract: Antigen-specific T cell immunity is HLA-restricted. Human immunodeficiency virus–type 1 (HIV-1) mutations that allow escape from host immune responses may therefore be HLA allele–specific. We analyzed HIV-1 reverse transcriptase sequences from a large HLA-diverse population of HIV-1–infected individuals. Polymorphisms in HIV-1 were most evident at sites of least functional or structural constraint and frequently were associated with particular host HLA class I alleles. Absence of polymorphism was also HLA allele–specific. At a population level, the degree of HLA-associated selection in viral sequence was predictive of viral load. These results support a fundamental role for HLA-restricted immune responses in driving and shaping HIV-1 evolution in vivo.

Analysis of total human immunodeficiency virus (HIV)-specific CD4(+) and CD8(+) T-cell responses: relationship to viral load in untreated HIV infection

Abstract: Human immunodeficiency virus (HIV)-specific T-cell responses are thought to play a key role in viral load decline during primary infection and in determining the subsequent viral load set point. The requirements for this effect are unknown, partly because comprehensive analysis of total HIV-specific CD4(+) and CD8(+) T-cell responses to all HIV-encoded epitopes has not been accomplished. To assess these responses, we used cytokine flow cytometry and overlapping peptide pools encompassing all products of the HIV-1 genome to study total HIV-specific T-cell responses in 23 highly active antiretroviral therapy naive HIV-infected patients. HIV-specific CD8(+) T-cell responses were detectable in all patients, ranging between 1.6 and 18.4% of total CD8(+) T cells. HIV-specific CD4(+) T-cell responses were present in 21 of 23 patients, although the responses were lower (0.2 to 2.94%). Contrary to previous reports, a positive correlation was identified between the plasma viral load and the total HIV-, Env-, and Nef-specific CD8(+) T-cell frequency. No correlation was found either between viral load and total or Gag-specific CD4(+) T-cell response or between the frequency of HIV-specific CD4(+) and CD8(+) T cells. These results suggest that overall frequencies of HIV-specific T cells are not the sole determinant of immune-mediated protection in HIV-infection.